Alzheimer's disease (AD) is a neurodegenerative illness characterized by memory loss and other cognitive deficits. McKhann et al., Neurology, 34: 939 (1984). It is the most common cause of dementia in the United States. AD can strike persons as young as 40-50 years of age, yet, because the presence of the disease is difficult to determine without dangerous brain biopsies, the time of onset is unknown. The prevalence of AD increases with age, with estimates of the affected population reaching as high as 40-50% by ages 85-90. Evans et al., JAMA, 262: 2551 (1989); Katzman, Neurology, 43: 13 (1993).
In practice, AD is definitively diagnosed through examination of brain tissue, usually at autopsy. Khachaturian, Arch. Neurol., 42: 1097 (1985); McKhann et al., Neurology, 34: 939 (1984). Neuropathologically, this disease is characterized by the presence of neuritic plaques (NP), neurofibrillary tangles (NFT) and neuronal loss along with a variety of other findings. Mann, Mech. Ageing Dev., 31: 213 (1985). Post-mortem slices of brain tissue of victims of Alzheimer's disease exhibit the presence of amyloid in the form of proteinaceous extracellular cores of the neuritic plaques that are characteristic of AD.
The amyloid cores of these neuritic plaques are composed of a protein called the β-amyloid (Aβ) that is arranged in a predominately beta-pleated sheet configuration. Mori et al., Journal of Biological Chemistry, 267: 17082 (1992); Kirschner et al., PNAS, 83: 503 (1986). Neuritic plaques are an early and invariant aspect of the disease. Mann et al., J. Neurol. Sci., 89: 169; Mann, Mech. Ageing Dev., 31: 213 (1985); Terry et al., J. Neuropathol. Exp. Neurol., 46: 262 (1987).
The initial deposition of Aβ probably occurs long before clinical symptoms are noticeable. The currently recommended “minimum microscopic criteria” for the diagnosis of AD is based on the number of neuritic plaques found in the brain. Khachaturian, Arch. Neurol., 42: 1097 (1985). Unfortunately, assessment of neuritic plaque counts must be delayed until after death.
Amyloid-containing neuritic plaques are a prominent feature of selective areas of the brain in AD as well as Down's Syndrome and in persons homozygous for the apolipoprotein E4 allele, who are very likely to develop AD. Corder et al., Science, 261: 921 (1993); Divry, P., J. Neurol. Psych., 27: 643 (1927); Wisniewski et al., Re-Examination of the Pathogenesis of the Senile Plaque: Progress in Neuropathology, (Grune and Stratton, N.Y. 1973), pp. 1-26. Brain amyloid is readily demonstrated by staining brain sections with thioflavin S or Congo red. Puchtler et al., J. Histochem Cytochem, 10: 35 (1962). Congo red stained amyloid is characterized by a dichroic appearance, exhibiting a yellow-green polarization color. The dichroic binding is the result of the beta-pleated sheet structure of the amyloid proteins. Glenner, G., N. Eng. J. Med., 302:1283 (1980). A detailed discussion of the biochemistry and histochemistry of amyloid can be found in Glenner, G., N. Eng. J. Med., 302: 1333 (1980).
Amyloidosis is a condition characterized by the accumulation of various insoluble, fibrillar proteins, generically termed “amyloid,” in the tissues of a patient. Amyloidosis is a slowly progressive condition, which can lead to significant morbidity and death. A diverse group of diseases or disease processes fall under the “amyloidosis” rubric, and can be termed amyloidosis-related diseases. An amyloid deposit is formed by the aggregation of amyloid proteins, followed by the further combination of aggregates and/or amyloid proteins. Formation and accumulation of aggregates of Aβ peptides in the brain are critical factors in the development and progression of AD. The fibrillar aggregates of amyloid peptides, Aβ1-40 and Aβ1-42, are major metabolic peptides derived from amyloid precursor proteins found in senile plaques and cerebrovascular amyloid deposits in AD patients. Xia et al., J. Proc. Natl. Acad Sci. U.S.A., 97: 9299 (2000). Prevention and reversal of Aβ plaque formation are being targeted as a treatment for this disease. Selkoe, D., JAAM, 283: 1615 (2000); Wolfe, M. S., et al., J. Med Chem., 41: 6 (1998); Skovronsky, D. M. et al., Trends Pharmacol. Sci., 21: 161 (2000).
In addition to the role of amyloid deposits in Alzheimer's disease, the presence of amyloid deposits has been shown in diseases such as glaucoma, Mediterranean fever, Muckle-Wells syndrome, idiopathic myeloma, amyloid polyneuropathy, amyloid cardiomyopathy, systemic senile amyloidosis, amyloid polyneuropathy, hereditary cerebral hemorrhage with amyloidosis, Down's syndrome, Scrapie, Creutzfeldt-Jacob disease, Kuru, Gerstmann-Straussler-Scheinker syndrome, medullary carcinoma of the thyroid, Isolated atrial amyloid, β2-microglobulin amyloid in dialysis patients, inclusion body myositis, β2-amyloid deposits in muscle wasting disease, Islets of Langerhans diabetes Type II insulinoma and other amyloidosis-related diseases.
Thus far, diagnosis of AD has been achieved mostly through clinical criteria evaluation, brain biopsies and post-mortem tissue studies. Research efforts to develop methods for diagnosing AD in vivo include (1) genetic testing, (2) immunoassay methods and (3) imaging techniques.
Evidence that abnormalities in Aβ metabolism are necessary and sufficient for the development of AD is based on the discovery of point mutations in the Aβ precursor protein in several rare families with an autosomal dominant form of AD. Hardy, Nature Genetics, 1: 233 (1992); Hardy et al., Science, 256: 184 (1992). These mutations occur near the N and C-terminal cleavage points necessary for the generation of Aβ from its precursor protein. St. George-Hyslop et al., Science, 235: 885 (1987); Kang et al., Nature, 325: 733 (1987). Genetic analysis of a large number of AD families has demonstrated, however, that AD is genetically heterogeneous. St. George-Hyslop et al., Nature, 347: 194 (1990). Linkage to chromosome 21 markers is shown in only some families with early-onset AD and in no families with late-onset AD. More recently, a gene on chromosome 14, whose product is predicted to contain multiple transmembrane domains and resembles an integral membrane protein, has been identified by Sherrington et al., Nature, 375: 754 (1995). This gene may account for up to 70% of early-onset autosomal dominant AD. Preliminary data suggests that this chromosome 14 mutation causes an increase in the production of Aβ. Scheuner et al., Soc. Neurosci. Abstr., 21: 1500 (1995). A mutation on a very similar gene has been identified on chromosome 1 in Volga German kindreds with early-onset AD. Levy-Lahad et al., Science, 269: 973 (1995).
Screening for apolipoprotein E genotype has been suggested as an aid in the diagnosis of AD. Scott, Nature, 366: 502 (1993); Roses, Ann. Neurol., 38: 6 (1995). Difficulties arise with this technology, however, because the apolipoprotein E4 allele is only a risk factor for AD, not a disease marker. It is absent in many AD patients and present in many non-demented elderly people. Bird, Ann. Neurol., 38: 2 (1995).
Immunoassay methods have been developed for detecting the presence of neurochemical markers in AD patients and to detect an AD related amyloid protein in cerebral spinal fluid. Warner, Anal. Chem., 59: 1203A (1987). These methods for diagnosing AD have not been proven to detect AD in all patients, particularly, at early stages of the disease, and are relatively invasive, requiring a spinal tap. Also, attempts have been made to develop monoclonal antibodies as probes for imaging of Aβ. Majocha et al., J. Nucl. Med., 33: 2184 (1992). The major disadvantage of antibody probes is the difficulty in getting these large molecules across the blood-brain barrier. Using antibodies for in vivo diagnosis of AD would require marked abnormalities in the blood-brain barrier in order to gain access into the brain. There is no convincing functional evidence that abnormalities in the blood-brain barrier reliably exist in AD. Kalaria, Cerebrovascular & Brain Metabolism Reviews, 4: 226 (1992).
Radiolabeled Aβ peptides have been used to label diffuse, compact and neuritic type plaques in sections of AD brain. However, these peptides share all of the disadvantages of antibodies. Specifically, peptides do not normally cross the blood-brain barrier in amounts necessary for imaging and because these probes react with diffuse plaques, they may not be specific for AD.
Data also suggest that amyloid binding compounds will have therapeutic potential in AD and type 2 diabetes mellitus. Morphological reactions including reactive astrocytosis, dystrophic neurites, activated microglia cells, synapse loss and full complement activation found around neuritic plaques all signify that neurotoxic and cell degenerative processes are occurring in the areas adjacent to these Aβ deposits. Joachim et al., Am. J. Pathol., 135: 309 (1989); Masliah et al., 137: 1293 (1990); Lue et al., Dementia, 3: 308 (1992). Aβ-induced neurotoxicity and cell degeneration has been reported in a number of cell types in vitro. Yankner et al., Science, 250: 279 (1990); Roher et al., BBRC, 174: 572 (1991); Frautschy et al., Proc. Natl. Acad. Sci., 88: 83362 (1991); Shearman et al., 91: 1470 (1994). It has been shown that aggregation of the Aβ peptide is necessary for in vitro neurotoxicity. Yankner, Neurobiol. Aging, 13: 615 (1992). Thus far, several laboratories have reported results which suggest that Congo red inhibits Aβ-induced neurotoxicity and cell degeneration in vitro. Burgevin et al., NeuroReport, 5: 2429 (1994); Lorenzo et al., Proc. Natl. Acad. Sci., 91: 12243 (1994); Pollack et al., Neuroscience Letters, 184: 113 (1995); Pollack et al., Neuroscience Letters, 197: 211 (1995). The mechanism appears to involve both inhibition of fibril formation and prevention of the neurotoxic properties of formed fibrils. Lorenzo et al., Proc. Natl. Acad. Sci., 91: 12243 (1994). Congo red also has been shown to protect pancreatic islet cells from the toxicity caused by amylin. Lorenzo et al., Proc. Natl. Acad. Sci., 91: 12243 (1994). Amylin is a fibrillar peptide similar to Aβ, which accumulates in the pancreas in type 2 diabetes mellitus.
The inability to assess amyloid deposition in AD until after death impedes the study of this devastating illness. Histological analysis of biopsy or autopsy materials also has its drawbacks. As such, a method of quantifying amyloid deposition before death is needed both as a diagnostic tool in mild or clinically confusing cases as well as in monitoring the effectiveness of therapies targeted at preventing Aβ deposition. Moreover, it remains of utmost importance to develop a safe and specific method for diagnosing AD before death by imaging amyloid in brain parenchyma in vivo. Even though various attempts have been made to diagnose AD in vivo, currently, there are few antemortem probes for brain amyloid. To date, simple, noninvasive methods for detecting and quantitating amyloid deposits in a patient have been eagerly sought. Thus, a need exists for amyloid binding compounds which enter the brain and bind selectively to amyloid. Similarly, amyloid binding compounds are also needed for detecting and quantitating amyloid deposits in other disease types including, for example, those previously mentioned. Amyloid binding compounds may also be needed for their therapeutic potential in the treatment and prophylaxis of, for example, AD and type 2 diabetes mellitus.